14-3-3ζ Regulates the Platelet Apoptosis During Storage

Yuzhong Yan; Rufeng Xie; Zhongping Ning

doi:10.1007/s12288-019-01229-z

14-3-3ζ Regulates the Platelet Apoptosis During Storage

Indian J Hematol Blood Transfus. 2020 Apr;36(2):324-329. doi: 10.1007/s12288-019-01229-z. Epub 2019 Nov 14.

Authors

Yuzhong Yan¹, Rufeng Xie², Zhongping Ning³

Affiliations

¹ 1Department of Transfusion, Shanghai University of Medicine and Health Sciences Affiliated Zhoupu Hospital, Shanghai, China.
² 2Blood Engineering Laboratory, Shanghai Blood Center, Shanghai, China.
³ 3Department of Cardiology, Shanghai University of Medicine and Health Sciences Affiliated Zhoupu Hospital, Shanghai, 201300 China.

Abstract

There is an increasing interest in the possibility of storing platelet concentrates below standard temperatures. The role of 14-3-3 proteins has been demonstrated in numerous cellular functions, including both its positive and negative roles in apoptosis. The 14-3-3ζ protein has a potential role in regulation of storage induced apoptosis in platelets. Apheresis platelets were collected and stored under either at room temperature (RT, 20-24 °C) or cold temperature (CT, 2-6 °C) conditions (n = 7 in each group). Flow cytometry was used to assess changes in phosphatidylserine and mitochondrial membrane potential in washed platelets. Proteomic changes were analyzed using Western blot and coimmunoprecipitation. During RT storage conditions used in this study, we found that both Annexin V and JC-1 exhibited significant increases at Day 3 compared to Day 1. In comparison to RT storage, a 3-day cold storage exhibited higher positive rates of Annexin V and lower positive rates of JC-1. The release of cytochrome c and caspase 3 and 9 cleavage were only observed in platelets maintained under RT storage conditions for 3 days. The anti-apoptosis protein Bcl-x_L was downregulated and the pro-apoptosis protein Bak was upregulated under RT storage conditions. However, both Bcl-x_L and Bak of CT-D3 exhibited no significant changes in comparison to either RT-D1 or CT-D1. Expression levels of 14-3-3ζ and GPIbα decreased significantly in RT-D3 compared to those in RT-D1, while the expression levels of CT-D3 were found to be significantly higher than those in RT-D3. Expression levels of Bad protein in CT-D3 were significantly lower than those in RT-D3. A comparative analysis of RT-D3 and CT-D3 demonstrated that both the ratios of Bad/14-3-3ζ and GPIbα/14-3-3ζ increased significantly following cold storage. The ratio was significantly larger following a 3-day cold storage in comparison to that at RT. During cold storage of platelets, the enhanced association between 14-3-3ζ and GPIbα was demonstrated to improve exposure of phosphatidylserine, and the enhanced association between 14-3-3ζ and Bad was shown to delay the depolarization of the mitochondrial membrane potential.

Keywords: 14–3–3ζ; Bad; Cold storage; GPIbα; Platelets.